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Genetically engineering encapsulin protein cage nanoparticle as a SCC-7 cell targeting optical nanoprobe.

Moon H, Lee J, Kim H, Heo S, Min J, Kang S - Biomater Res (2014)

Bottom Line: Encapsulin protein cage nanoparticle is used to develop a cell-specific targeting optical nanoprobe.FcBPs are genetically inserted and successfully displayed on the surface of encapsulin to form FcBP-encapsulin.Encapsulin protein cage nanoparticle is robust enough to maintain their structure at high temperature and easily acquires multifunctions on demand through the combination of genetic and chemical modifications.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, School of Life Sciences, Ulsan National Institute of Science and Technology (UNIST), Ulsan, 689-798 South Korea.

ABSTRACT

Background: Protein cage nanoparticles are promising nanoplatform candidates for efficient delivery systems of diagnostics and/or therapeutics because of their uniform size and structure as well as high biocompatibility and biodegradability. Encapsulin protein cage nanoparticle is used to develop a cell-specific targeting optical nanoprobe.

Results: FcBPs are genetically inserted and successfully displayed on the surface of encapsulin to form FcBP-encapsulin. Selectively binding of FcBP-encapsulin to SCC-7 is visualized with fluorescent microscopy.

Conclusions: Encapsulin protein cage nanoparticle is robust enough to maintain their structure at high temperature and easily acquires multifunctions on demand through the combination of genetic and chemical modifications.

No MeSH data available.


Binding behaviors of FcBP-Encapsulin to antibodies. (A) QCM resonance frequency change (-ΔF) profiles of either encapsulin (dashed line) or FcBP-encapsulin (solid line) on the gold QCM sensors and subsequent deposition of rabbit IgG on the monolayer of FcBP-encapsulin. Filled and open arrows indicate the timing of encapsulin variant and rabbit IgG introductions, respectively. Thin arrows indicate the timing of buffer washing. SPR analyses of FcBP-encapsulin (B) and encapsulin (C) bindings to rabbit IgG immobilized gold SPR sensors. Filled and open arrows indicate the timing of encapsulin variant introductions and buffer washing, respectively. Inset of (C) is amplified graph of low RU ranges.
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Fig2: Binding behaviors of FcBP-Encapsulin to antibodies. (A) QCM resonance frequency change (-ΔF) profiles of either encapsulin (dashed line) or FcBP-encapsulin (solid line) on the gold QCM sensors and subsequent deposition of rabbit IgG on the monolayer of FcBP-encapsulin. Filled and open arrows indicate the timing of encapsulin variant and rabbit IgG introductions, respectively. Thin arrows indicate the timing of buffer washing. SPR analyses of FcBP-encapsulin (B) and encapsulin (C) bindings to rabbit IgG immobilized gold SPR sensors. Filled and open arrows indicate the timing of encapsulin variant introductions and buffer washing, respectively. Inset of (C) is amplified graph of low RU ranges.

Mentions: A synthetic cyclic FcBP (DCAWHLGELVWCT) exhibited a high affinity to rabbit IgGs with the value of Kd =305 nM [32]. In order to test whether inserted FcBPs are on the surface of encapsulin, we performed real-time quartz crystal microbalance (QCM) analysis. Typically, deposition of molecules on the QCM sensor results in decreases in resonance frequency (-ΔF) and the extent of frequency changes are sensitive to the masses of the deposited molecules [30, 34]. We have shown that QCM analysis is a nice method to determine protein-protein interactions. Various types of protein cages, including ferritins, lumazine synthase, and virus-like particles (VLP), showed strong binding capability to the gold QCM sensor and form uniform monolayers without any surface modifications [30, 34]. The resonance frequency value of FcBP-encapsulin (Figure 2A, solid line, filled arrow) itself was lower than that of encapsulin (Figure 2A, dashed line, filled arrow) only probably due to the increase in mass of the FcBP insertions to encapsulin (24 amino acids, an approximate 8% increase in mass) (Figure 2A). The resonance frequency of each sample decreased, plateauing when it reached a certain value. In addition, no additional changes were observed even with continuous introduction of further sample and subsequent washing, indicating that the encapsulin had formed a uniform monolayer regardless of the FcBP insertion and that there was negligible amount of non-specific absorption (Figure 2A, thin arrow). Subsequently, we introduced rabbit IgG solution over the encapsulin- or FcBP-encapsulin-monolayered QCM sensors and measured frequency changes in real-time (Figure 2A). While the frequency of FcBP-encapsulin-monolayered QCM sensor decreased dramatically upon introduction of rabbit IgG, the frequency of encapsulin-monolayered QCM sensor remained unchanged (Figure 2A, open arrow). Extensive washing removed only slight amounts of non-specifically associated rabbit IgG with most of the initially bound rabbit IgG remaining bound (Figure 2A, thin arrow). These results indicated that the inserted FcBPs are well displayed on the surface of encapsulin and they are fully accessible for Fc regions of rabbit IgGs to bind. These data also imply that exterior displayed FcBPs can be used as ligands for target selective delivery of FcBP-encapsulin.Figure 2


Genetically engineering encapsulin protein cage nanoparticle as a SCC-7 cell targeting optical nanoprobe.

Moon H, Lee J, Kim H, Heo S, Min J, Kang S - Biomater Res (2014)

Binding behaviors of FcBP-Encapsulin to antibodies. (A) QCM resonance frequency change (-ΔF) profiles of either encapsulin (dashed line) or FcBP-encapsulin (solid line) on the gold QCM sensors and subsequent deposition of rabbit IgG on the monolayer of FcBP-encapsulin. Filled and open arrows indicate the timing of encapsulin variant and rabbit IgG introductions, respectively. Thin arrows indicate the timing of buffer washing. SPR analyses of FcBP-encapsulin (B) and encapsulin (C) bindings to rabbit IgG immobilized gold SPR sensors. Filled and open arrows indicate the timing of encapsulin variant introductions and buffer washing, respectively. Inset of (C) is amplified graph of low RU ranges.
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Related In: Results  -  Collection

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Fig2: Binding behaviors of FcBP-Encapsulin to antibodies. (A) QCM resonance frequency change (-ΔF) profiles of either encapsulin (dashed line) or FcBP-encapsulin (solid line) on the gold QCM sensors and subsequent deposition of rabbit IgG on the monolayer of FcBP-encapsulin. Filled and open arrows indicate the timing of encapsulin variant and rabbit IgG introductions, respectively. Thin arrows indicate the timing of buffer washing. SPR analyses of FcBP-encapsulin (B) and encapsulin (C) bindings to rabbit IgG immobilized gold SPR sensors. Filled and open arrows indicate the timing of encapsulin variant introductions and buffer washing, respectively. Inset of (C) is amplified graph of low RU ranges.
Mentions: A synthetic cyclic FcBP (DCAWHLGELVWCT) exhibited a high affinity to rabbit IgGs with the value of Kd =305 nM [32]. In order to test whether inserted FcBPs are on the surface of encapsulin, we performed real-time quartz crystal microbalance (QCM) analysis. Typically, deposition of molecules on the QCM sensor results in decreases in resonance frequency (-ΔF) and the extent of frequency changes are sensitive to the masses of the deposited molecules [30, 34]. We have shown that QCM analysis is a nice method to determine protein-protein interactions. Various types of protein cages, including ferritins, lumazine synthase, and virus-like particles (VLP), showed strong binding capability to the gold QCM sensor and form uniform monolayers without any surface modifications [30, 34]. The resonance frequency value of FcBP-encapsulin (Figure 2A, solid line, filled arrow) itself was lower than that of encapsulin (Figure 2A, dashed line, filled arrow) only probably due to the increase in mass of the FcBP insertions to encapsulin (24 amino acids, an approximate 8% increase in mass) (Figure 2A). The resonance frequency of each sample decreased, plateauing when it reached a certain value. In addition, no additional changes were observed even with continuous introduction of further sample and subsequent washing, indicating that the encapsulin had formed a uniform monolayer regardless of the FcBP insertion and that there was negligible amount of non-specific absorption (Figure 2A, thin arrow). Subsequently, we introduced rabbit IgG solution over the encapsulin- or FcBP-encapsulin-monolayered QCM sensors and measured frequency changes in real-time (Figure 2A). While the frequency of FcBP-encapsulin-monolayered QCM sensor decreased dramatically upon introduction of rabbit IgG, the frequency of encapsulin-monolayered QCM sensor remained unchanged (Figure 2A, open arrow). Extensive washing removed only slight amounts of non-specifically associated rabbit IgG with most of the initially bound rabbit IgG remaining bound (Figure 2A, thin arrow). These results indicated that the inserted FcBPs are well displayed on the surface of encapsulin and they are fully accessible for Fc regions of rabbit IgGs to bind. These data also imply that exterior displayed FcBPs can be used as ligands for target selective delivery of FcBP-encapsulin.Figure 2

Bottom Line: Encapsulin protein cage nanoparticle is used to develop a cell-specific targeting optical nanoprobe.FcBPs are genetically inserted and successfully displayed on the surface of encapsulin to form FcBP-encapsulin.Encapsulin protein cage nanoparticle is robust enough to maintain their structure at high temperature and easily acquires multifunctions on demand through the combination of genetic and chemical modifications.

View Article: PubMed Central - PubMed

Affiliation: Department of Biological Sciences, School of Life Sciences, Ulsan National Institute of Science and Technology (UNIST), Ulsan, 689-798 South Korea.

ABSTRACT

Background: Protein cage nanoparticles are promising nanoplatform candidates for efficient delivery systems of diagnostics and/or therapeutics because of their uniform size and structure as well as high biocompatibility and biodegradability. Encapsulin protein cage nanoparticle is used to develop a cell-specific targeting optical nanoprobe.

Results: FcBPs are genetically inserted and successfully displayed on the surface of encapsulin to form FcBP-encapsulin. Selectively binding of FcBP-encapsulin to SCC-7 is visualized with fluorescent microscopy.

Conclusions: Encapsulin protein cage nanoparticle is robust enough to maintain their structure at high temperature and easily acquires multifunctions on demand through the combination of genetic and chemical modifications.

No MeSH data available.